PROJECT SUMMARY Increasing evidence has supported a role for chromatin misregulation in the pathogenesis of various human diseases including cancer, although the underlying mechanism remains unclear. Recent tumor genome sequencing efforts have identified inactivating mutations in NSD1, a histone H3 lysine 36 (H3K36)-specific methyltransferase, in 10-15% of head and neck squamous cell carcinomas (HNSCC), the sixth leading cancer worldwide. This study focuses on elucidating the molecular mechanisms by which NSD1 inactivation facilitate HNSCC development and will test the central hypothesis that biochemical and genetic inactivation of NSD1 alters DNA methylome and transcriptome to disrupt squamous tissue differentiation and promote HNSCC initiation in vivo. During the mentored training period, I will characterize the biochemical basis and functional importance of NSD1 inhibition by H3 lysine 36 to methionine (H3K36M) mutation, an oncohistone previously associated with pediatric malignancies and identified in HNSCC through my preliminary studies. I will also profile the impact of NSD1 and H3K36M mutations on the genome-wide distribution of H3K36 methylation and DNA methylation as well as on the transcriptome. During the independent phase of the award period, I propose to investigate the function of NSD1 in squamous tissue differentiation and in vivo suppression of HNSCC. Together the proposed studies will provide novel insights into the pathogenesis of a subset of genetically defined and epigenetically distinct HNSCC that can translate into improved care for patients. They will also serve as a platform for me to obtain trainings in chemical biology, computational epigenomics and mouse models of cancer that will critically contribute to my career development as an independent investigator in the field of cancer epigenetics.